Ions may be generated from relatively heavy analyte molecules, having molecular weights of several hundred to several thousand Daltons, using an electrospray ion source. For example, a relatively high voltage (e.g., several kilovolts) may be applied to a pointed spray capillary, containing spray liquid with dissolved analyte molecules, to generate a relatively strong electric field around the tip. The electric field polarizes and charges the surface of the spray liquid in the open tip. In this manner, an electric tractive force creates a so-called “Taylor cone” on the surface of the liquid, and an electric field draws a fine jet of liquid out of the tip of the cone. The jet is intrinsically unstable due to its high surface charge, which opposes the surface tension. The jet disintegrates by constriction into minute, highly charged droplets with diameters on the order of a hundred nanometers to a few micrometers.
The size of the droplets depends on the tip aperture and the electric field generated around the tip of the spray capillary. Nanoelectrospray ionization, for example, uses apertures of approximately two to three micrometers. Such tip apertures can generate droplets with 100 to 200 nanometer diameters using a spray voltage of less than one kilovolt at flow rates of a few tens to a maximum of one thousand nanoliters per minute. “Normal” electrospraying, on the other hand, uses tip apertures of approximately ten to thirty micrometers in diameter. Such tip apertures can generate droplets with one to two micrometer diameters using a spray voltage of three to five kilovolts at flow rates of one to a several thousand microliters per minute. The droplets generated with normal electrospraying therefore have a volume more than a thousand times larger than nanospray droplets. Droplets with a diameter of around one micrometer carry about 50,000 elementary charges.
The decomposition of the jet of liquid into droplets can be supplemented using a focused jet of spray gas. The spray gas is blown in around the tip of the capillary by a concentric spray gas capillary. This causes the jet of droplets to be guided in a somewhat more focused manner, although the droplets are created with greater diameter variance.
The droplets are evaporated in a hot drying gas to vaporize neutral solvent molecules. This causes the charge density on the surface to continuously increase. When the charge density on the surface becomes so large that the Coulomb repulsion exceeds a force of cohesive surface tension (i.e., “Rayleigh limit”), the droplets split into smaller droplets. The unstable surface brings about random oscillations of the fluid on the surface, for example, cause the droplets to split. The separation of the droplets, however, causes the charges of both droplets to fall below the Rayleigh limit.
The smaller droplets have a higher charge relative to their mass since the total charge qR of a droplet at the Rayleigh limit is proportional to the root of the third power of the diameter d, (qR≈√d3). Each small droplet therefore has, for example, two per cent of the mass, but fifteen per cent of the charge. Both small and large droplets, however, have a mass-to-charge ratio above the Rayleigh limit and thus continue to vaporize.
Different-sized droplets, with charge densities at their respective Rayleigh limit, have different electric mobilities μ=v/E (v=velocity) when being pulled through gas by electric fields of field strength “E”. Slow motions of the droplets without eddying are subject to Stokes' law, exhibiting friction proportional to the diameter “d” and the velocity “v” of the droplets. The electric mobility “μ” is therefore proportional to √d. As a result, larger droplets have higher electric mobilities than smaller droplets. Rapid motions with turbulent eddying are subject to Newtonian friction, proportional to the cross-section d2 and the square of the velocity v2. Under these conditions, which usually do not, however, apply to spray droplets and their velocities in drying gases, the electric mobility μ is proportional to the inverse root E−2 of the field strength E and the inverse fourth root d−4 of the diameter d.
Each droplet, large and small, continues to vaporize. The rate of vaporization of the small droplets increases since the coordination number of their surface molecules gets smaller. The vapor pressure therefore increases until the splitting off and vaporization processes end relatively rapidly in the complete drying of the droplets leaving multiply charged analyte ions that were included in the droplets. In the last phase, protonated water molecules may also vaporize. The fowled analyte ions are, for example, generally only surrounded by a somewhat more strongly bound sheath of one to two molecular layers of solvent molecules, usually water molecules.
As indicated above, there are two basic types of electrospraying: nanoelectrospraying (also referred to as “nanospraying”) and normal electrospraying (also referred to as “microspraying”). In nanospraying, the droplets are typically sprayed directly into the inlet capillary leading to the ion analyzer. In microspraying, in contrast, analyte ions generated by being vaporized in free air are directed into the inlet capillary.
Nanospraying generates very small droplets with substantially equal diameters (e.g., approximately 100 to 200 nanometers). The droplets are drawn into the inlet capillary by and accelerated in a transport gas. A typical transport gas includes nitrogen at a temperature between room temperature and 300 degrees Celsius. The droplets vaporize slowly in the inlet capillary, supported by an ever decreasing pressure.
Advantageously, gas-dynamic focusing can guide the droplets through the capillary with relatively low losses. In the inlet capillary, for example, brief boundary turbulences give way to a stable laminar flow of the transport gas with a parabolic velocity profile; i.e., where the flow is fastest in the center and resting at the capillary wall. The droplets are entrained by the fast gas stream, and are held in the axis of the capillary. When droplets swerve to the side, they enter a gas stream region which flows at different speeds on either side. The Bernoulli effect drives the particles back toward the axis using lift, similar to that experienced by an airplane wing.
The greater the difference in velocity between the gas and droplets, the stronger the “Bernoulli focusing”. This is because the lift is proportional to the difference between the squares of the velocities on either side of the droplet. Since the gas flow rate increases towards the end of the inlet capillary, the droplets do not travel as fast as the gas. The focusing effect is therefore maintained until the droplets are completely vaporized or have left the capillary. An opposing electric field in the inlet capillary can further enhance the focusing effect by decelerating the droplets to prevent them from assuming the velocity of the gas. It is unclear, however, whether larger analyte ions are also subject to the Bernoulli focusing when they are decelerated by an opposing field, and whether this focusing can be effective against the repulsion by the space charge prevailing in the axis.
Disadvantageously, the spray capillary tips used for nanospraying must be precisely adjusted with respect to the inlet capillary aperture. The ion sources therefore are usually equipped with microscopes or micro cameras for aligning the tips. Furthermore, as indicated above, nanospraying has relatively low flow rates.
During microspraying, in contrast, the droplets are vaporized outside the inlet capillary, thus releasing the analyte ions. The analyte ions generally have a solvate layer. The analyte ions lose the solvate layer, however, on their way through the inlet capillary into the vacuum system as they are heated by hot transport gas and as pressure along the inlet capillary decreases. The hot transport gas directs the analyte ions into the admission aperture of the inlet capillary. However, very few of the analyte ions are directed into the admission aperture due to the large volume of analyte ions that are produced. Microspraying therefore feeds far less than one percent of the sprayed analyte molecules into the ion analyzer in ionized form.
During both nanospraying and microspraying, the inlet capillary guides the charged analyte molecules into a vacuum system of an ion analyzer. The ion analyzer can be, for example, a mass spectrometer or an ion mobility spectrometer. The analyte ions are captured in the vacuum system by, for example, an ion funnel separated from the accompanying transport gas and introduced into the ion analyzer via additional ion guides and pumping stages. The analyte ions are analyzed in the ion analyzer. A single inlet capillary can be used to introduce the analyte ions into the vacuum. Several inlet capillaries can also be bundled together to introduce the analyte ions into the vacuum. Hereinafter the term “inlet capillary” shall be used generically to refer to both a sole capillary and a bundle of capillaries.
Most of the analyte ions are multiply charged. The number of charges, however, can vary for a single substance. The average number of charges increases roughly in proportion to the mass of the analyte ions. For heavy ions, for example, the mass-to-charge ratios m/z (m=mass; z=number of excess elementary charges of the ion) have a wide distribution from approximately m/z=700 Daltons to approximately m/z=1,600 Daltons. The heavy molecules of albumin (m=66 kDa), for example, have a 50-fold charge on average, while light molecules with molecular weights below m=2 kDa are predominantly singly charged. The distribution of the charges can be affected by the composition of the solvent, the spraying processes and the processes with which the ions are guided through gases.
The droplets in the jet of spray strongly repel each other since each droplet is highly charged (e.g., with 50,000 elementary charges for one droplet with a diameter of one micrometer). The jet of spray droplets accelerated in the electric field therefore broaden to a cloud with a pronounced funnel shape almost immediately after the droplets have been formed. During nanospraying, the broadening is limited by the transport gas with which the cloud of droplets is drawn into the inlet capillary and which entrains and accelerates the droplets. During microspraying, in contrast, the volume of the gas containing the analyte ions after the liquid has vaporized from the droplets is considerably larger. It is therefore difficult to draw a large number of analyte ions from the large volume into the inlet capillary. A focused jet of spray gas, which can be heated up to about 150° C., can be introduced concentrically around the spray capillary to reduce the volume in a radial direction. The reduction in volume, however, further accelerates the spray droplets. This produces an elongated ion formation volume of moderate width, but in which many fast, unvaporized droplets are flying through the cloud of analyte ions.
An elongated and moderately wide volume with analyte ions is produced from the spray gas. The ions are usually extracted perpendicularly and fed to the inlet capillary. The extraction is generally successful for only a small fraction of the analyte ions, however, because only analyte ions from a small section of the length and width of the ion formation volume reach the inlet capillary. A “super hot” sheath gas at a temperature of about 300° C. is blown in around the hot spray gas to focus the ion formation volume in the radial direction and, therefore, “thermally focus” the droplets.
Although normal electrospraying ionizes most of the analyte molecules when the droplets are completely vaporized, the yield of ions introduced into the analyzer is relatively small. Nevertheless, normal electrospraying is widely used because it can be easily coupled to the normal flow rates of analytical liquid chromatography (HPLC). Nanospraying, in contrast, provides a relatively high yield of analyte ions. Nanospraying, however, cannot typically be coupled with liquid chromatography without splitting the flow of liquid since nano-HPLC has flow rates which are far above those which nanospraying can cope with. The unfavorable splitting of the liquid flow therefore cancels out the favorable ion yield. Attempts to inject larger droplets, which are produced at slightly higher flow rates with slightly larger spray tip aperture diameters, directly into the inlet capillary have so far been unsuccessful.
There is a need for an improved technique of guiding spray droplets into an inlet capillary of a mass spectrometer.